Lipoprotein lipase (LPL) is a secretory glycoprotein which contributes to the delivery of lipid fuels to tissues where they are predominantly stored (adipose tissue) or oxidized (muscle). The regulation of LPL in adipose tissue and muscle is often divergent, and alterations in LPL and its regulation by hormones and nutrients are commonly seen in metabolic disorders of insulin resistance, i.e. obesity and type II diabetes mellitus. The importance of LPL in muscle to the regulation of energy balance has recently been demonstrated by the prevention of high fat diet-induced obesity in mice with targeted muscle- specific overexpression of LPL (MCKhLPL). Of interest, despite the reduction in plasma free fatty acids and triglycerides in MCKhLPL mice, muscle triglycerides were increased and insulin sensitivity decreased. This raises important questions about the contribution of LPL to fatty acid metabolism in muscle. In the current application, a series of related studies will be implemented to pursue the mechanisms by which LPL overexpression in muscle favorably alters body composition, but results in triglyceride accumulation in muscle and reductions in insulin sensitivity. In addition, a combined in vivo and in vitro approach will be used. Investigations will focus on the following hypotheses, that: 1) deprivation of energy intake, reductions in ambient temperature and exercise will modify energy expenditure and reduce adipose tissue mass more in MCKhLPL mice than controls, and that these perturbations will also affect the phenotype of female MCKhLPL mice, previously unaffected by LPL overexpression in skeletal muscle; and 2) in mice and in cultured C2C12 myoblasts stably transfected with LPL, that LPL will partition lipoprotein fatty acids preferentially to pathways of esterification prior to oxidation. Overall, these studies should provide new and important insights into the physiology of LPL in muscle, and how enhancing LPL expression in muscle affects fatty acid and triglyceride metabolism in muscle, insulin action and body composition.